Instruction Manual OmniPAGE Vertical

Instruction Manual OmniPAGE Vertical

 

Instruction Manual

OmniPAGE Vertical Electrophoresis Units

Catalogue Numbers

CVS10D, CVS10DSYS, CVS10PRE

VS10D, VS10DSYS, VS10PRE, VS10DCAST

VS10WD, VS10WDSYS

VS20D, VS20DSYS, VS20DCAST,

VS30D, VS30DSYS, VS30DCAST

1  

 

Contents

Section 1 General Information

1.1

1.2

1.3

1.4

Safety Instructions

Introduction

Product Information

Packing List

1.5

1.6

1.4.1

1.4.2

1.4.3

1.4.4

omniPAGE Mini omniPAGE Mini Wide omniPAGE Maxi omniPAGE VS30

Specifications

Care and Maintenance

Section 2 Operating Instructions

2.1 Setting up the omniPAGE Gel Tank

2.2 Gel casting for CVS10, VS20W, VS20 and VS30

2.2.1) Cleaning the Glass Plates

2.2.2) Glass cassette Assembly

2.2.3) Casting Stand Assembly

2.2.4) Gel Pouring

2.3) o mniPAGE module asse mbly and Sample loading

2.4) Gel Running

2.5) Gel Staining

Section 3 Gel Preparatio n

3.1 Gel Selection

3.2 Volumes required per gel

3.3 Gel Preparation

3.4 Stacking Gel Preparation

3.5 Buffer Volume

3.6 Gel Running Conditions

3.7 Preparation of denatured pr otein samples for loading

Sectio n 4 Stock Solutions

Sectio n 5 Ordering Information

5.1 omniPAGE Mini

5.2 omniPAGE Mini Wide

5.3 omniPAGE Maxi

5.4 omniPAGE VS30

Sectio n 6 Warranty

2  

Page

23

23

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26

29

31

20

20

21

22

17

18

1 9

12

14

16

16

16

11

11

11

12

5

6

7

8

9

10

2

3

4

5

Section 1

General In formation

1.1) Safety Instructio ns

 

WHEN USED CORRECTLY, THESE UNITS POSE NO HEALTH RISK.

HOWEVER, THESE UNITS CAN DELIVER DANGEROUS LEVELS OF

ELECTRICITY AND ARE TO BE OPERATED ONLY BY QUALIFIED

PERSONNEL FOLLOWING THE GUIDELINES LAID OUT IN THIS

INSTRUCTION MANUAL.

ANYONE INTENDING TO U SE THIS EQUIPMENT SHOULD READ THE

COMPLETE MANUAL THOROUGHLY.

THE UNIT MUST NEVER BE USED WIT HOUT THE SAFETY LID

CORRECTLY IN POSITION.

THE UNIT SHOULD NOT BE USED IF THERE IS ANY SIGN OF

DAMAGE TO THE EXTERNAL TANK OR LID.

ACRYLAMIDE IS A POWERFUL NEUROTOXIN IN SOLUTION FORM.

POLYMERIZED GELS CAN CONTAIN SOME UNPOLYMERIZED

SOLUTION AND PROTECTIVE GLOVES AND CLOTHING MUST B E

WORN.

THESE UN ITS COMPLY WITH THE STATUTORY CE SAFETY

DIRECTIVES: 73/23/EEC: LOW VOLTAGE DIRECTIVE: IEC 1 010-

1:1990 plus AMENDMENT 1:1992

EN 61010-1:1993/BS EN 61010-1:1993

3  

1.2) Introduction

Cleaver Scientifics’ omniPAGE range of Vertical Gel Units combines ease of use with high resolution separations. Four sizes, Mini 10 x 10cm, Mini Wide

20 x 10cm and Maxi 20 x 20cm, VS 30 x 30cm share a host of common features including a guaranteed leak proof seal required for trouble free, rapid and uncomplicated gel casting. Utilising a built in gel running module eliminates time consuming transfer of glass plates during casting, a process which can cause gel damage and misalignment. Glass plates with permanently bonded spacers guarantee perfect spacer alignment. The glass plate sandwich is then simply inserted between pressure bars and the new zero screw slide clamps clicked into position. This ensures fast set up times while even pressure bars and ultra soft seals guarantee leak proof casting.

Once the gel has polymerised, the gel running module is just inserted into the gel tank for electrophoresis.

 

4  

 

1.3) Product Information

Cleaver Scientifics’ vertical electrophoresis units comes as a complete package including Gel Tank, Safety Lid, Gel Casting Inner Module, Glass plates with bonded spacers, Combs, Multipurpose Key, Power Cables and

Cooling pads. Detailed description of all the components is written below.

Gel Tank and

Safety Lid

Inner Module

Glass Plates

Multipurpose Key

Power Cables

Cooling Pads

Combs with special gel loading guides.

-Injection moulded gel tank and safety lid provides a sealed electrophoresis system which is compatible with all major types of 8 X 10cm and 10 x 10cm precast gel. All omniPAGE tanks are equipped with specially designed thumb locators for lifting up the safety lid easily.

-All safety lids are designed to accommodate the polarity design of the unit. They have special tapped holes which connect with the leading edge of the power cables.

Injection moulded inner module gives a twofold effectiveness for gel casting and running, thus no need of transferring the glass plates after casting the gel.

2mm thick glass plates for the omniPAGE mini and 4mm thick glass plates for rest of the range prevents breakage and have bonded spacers for convenience. All our spacers are colour coded depending upon their thickness.

CVS10KEY or the multipurpose key can be easily used to separate your notched and plain glass plate to release the gel.

The same key can be used to open the CVS10 clamping doors.

Note: Not for CVS20 and CVS30.

Cleaver Scientifics’ power cables are designed with protective retractable connectors which are compatible with most power supplies.

Rapid set up cooling packs enhance resolution eliminating the need of a chiller.

Special combs also known as the Combicombs are designed with special oval shaped gel loading guides on the other end of an ordinary comb which can be used

5  

1.4) Packing List

The packing lists should be referred to as soon as the units are received to ensure that all components have been included. The unit should be checked for damage when received. Please contact your supplier if there are any problems or missing items.

1.4.1) omniPAGE Mini

CVS10D, CVS10DSYS, CVS10PRE,

VS10D, VS10DSYS, VS10PRE, VS10DCAST

Each unit includes a tank, lid, internal module, electrodes and the following accessories:-

Glass Plates Combs

VS10D

CVS10D

Casting base Cooling

Pack

Cables SCREWS

VS10ICB CSL-CAB VS10-

SCREW x 4

VS10DSYS

CVS10DSYS

VS10PRE

CVS10PRE

VS10DCAST

VS10NG –

Notched, Pk/2

VS10PGS1 –

Plain with bonded 1mm spacers, Pk/2

VS10-DP –

Dummy Plate

VS10NG –

Notched, Pk/2

VS10PGS1 –

Plain with bonded 1mm spacers, Pk/2

VS10-DP –

Dummy Plate

VS10-DP –

Dummy Plate

2 of VS10-

12-1

1mm thick,

12 sample

2 of VS10-

12-1

1mm thick,

12 sample

VS10DCAST

VS10DCASTM

– Mat

VS10ICB CSL-CAB VS10-

SCREW x 4

SCREW x 4

VS10DCAST

VS10DCASTM

- Mat

6  

 

1.4.3) omniPAGE Mini Wide

VS10WD, VS10WDSYS

Each unit includes a tank, lid, internal module, electrodes and the following accessories:-

Glass Plates Combs Casting base Cooling

Pack

Cables

VS10WD

VS10WDSYS

VS10WNG - Notched,

Pk/2

VS10WPGS1 – Plain with bonded 1mm spacers, Pk/2

VS10W-DP – Dummy

Plate

VS10WNG - Notched,

Pk/2

VS10WPGS1 – Plain with bonded 1mm spacers, Pk/2

VS10W-DP – Dummy

Plate

2 of VS20-24-1

1mm thick, 24 sample

2 of VS20-24-1

1mm thick, 24 sample

VS20DCAST

VS20DCASTM -

Mat

VS20ICB CSL-CAB

VS20DCAST

VS20DCAST

VS20DCASTM -

Mat

1.4.3) omniPAGE Maxi

VS20D, VS20DSYS, VS20DCAST

Each unit includes a tank, lid, internal module, electrodes and the following accessories:-

Cables

VS20D

Glass Plates Combs

VS20NG - Notched,

Pk/2

VS20PGS1 – Plain with bonded 1mm spacers,

Pk/2

VS20-DP – Dummy

Plate

2 of VS20-24-1

1mm thick, 24 sample

Casting base Cooling

Pack

7  

 

VS20DSYS

VS20NG - Notched,

Pk/2

VS20PGS1 – Plain with bonded 1mm spacers,

Pk/2

VS20-DP – Dummy

Plate

2 of VS20-24-1

1mm thick, 24 sample

VS20DCAST

VS20DCASTM -

Mat

VS20ICB CSL-CAB

VS20DCAST

VS20DCAST

VS20DCASTM -

Mat

1.4.5) omniPAGE VS30

VS30D, VS30DSYS, VS30DCAST

Each unit includes a tank, lid, internal module, electrodes and the following accessories:-

Glass Plates Combs Casting base Cooling

Pack

Cables

VS30D

VS30DSYS

VS30DCAST

VS30NG - Notched,

Pk/2

VS30PGS1.5 – Plain with bonded 1.5mm spacers, Pk/2

VS30-DP – Dummy

Plate

VS30NG - Notched,

Pk/2

VS30PGS1.5 – Plain with bonded 1.5mm spacers, Pk/2

VS30-DP – Dummy

Plate

2 of VS30-1-

1.5

1.5mm thick,

1 sample

2 of VS30-1-

1.5

1.5mm thick,

1 sample

VS30DCAST

VS30DCASTM -

Mat

VS30DCAST

VS30DCASTM -

Mat

VS30ICB CSL-CAB

8  

 

1.5) Specifications

Plate

Dimensions

Gel Dimensions

(WxL)

omniPAGE

Mini

10x10cm

7.5x8cm

omniPAGE

Mini Wide

20 X 10cm

18 X 8cm

Unit

Dimensions

Max Sample

Capacity

Buffer Volume

Combs Available

No. of Teeth

Thickness

Environmental

Operating

Conditions

omniPAGE

Maxi

20 X 20CM

16 X 17.5CM

omniPAGE

VS30

30 X 22cm

28 X 20cm

19x13x15cm

(W x D x H)

80 Samples

20 Samples per Gel

26 X 16 X 16cm

(W X D X H)

192 Samples

48 Samples per Gel

26 X 16 X 28CM

(W X D X H)

192 Samples

48 Samples per Gel

36 X 33 X 18CM

(W X H X D)

300 Samples per

Run

75 Samples per Gel

Min1800ml

Max 8400ml

Min 250ml,

Max 1200ml

Min 600ml

Max 2800ml

Min 1200ml

Max 5600ml

1, 5, 8MC, 9, 10,

12, 16MC, 20

0.75, 1, 1.5, 2mm

1, 5, 10, 18MC, 24,

30, 36MC, 48

0.75, 1, 1.5, 2mm

1, 5, 10, 18MC, 24,

30, 36MC, 48

0.75, 1, 1.5, 2mm

1, 2, 4, 28MC,

56MC, 75

0.25, 0.35, 0.5, 1,

1.5, 2.0mm

Maximum Altitude

2,000 m

Temperature

Range 4°C - 65°C

Humidity

Upto 80%

Not for outdoor

Use

Maximum Altitude

2,000 m

Maximum Altitude

2,000 m

Temperature

Range 4°C - 65°C

Humidity

Upto 80%

Not for outdoor

Use

Temperature

Range 4°C - 65°C

Humidity

Upto 80%

Not for outdoor

Use

Maximum Altitude

2,000 m

Temperature

Range 4°C - 65°C

Humidity

Upto 80%

Not for outdoor

Use

This apparatus is rated POLLUTION DEGREE 2 in accordance with IEC 664.

POLLUTION DEGREE 2, states that: “Normally only non-conductive pollution occurs.

Occasionally, however, a temporary conductivity caused by condensation must be expected”.

9  

 

 

1.6) Care and Maintenance

Cleaning Large Format Vertical Units

Units are best cleaned using warm water and a mild detergent.

Water at temperatures above 60

0

C can cause damage to the unit and

components.

The inner module should be thoroughly rinsed with warm water or distilled water to prevent build up of salts but care should be taken not to damage the enclosed electrode and vigorous cleaning is not necessary or advised. Air drying preferably before use.

The units should only be cleaned with the following:-

Warm water with a mild concentration of soap or other mild detergent.

Compatible detergents include dishwashing liquid, Hexane and

Aliphatic hydrocarbons. The units should not be left in detergents for more than 30 minutes.

The units should never come into contact with the following cleaning agents, these will cause irreversible and accumulative damage:-

Acetone, Phenol, Chloroform, Carbon tetrachloride, Methanol,

Ethanol, Isopropyl alcohol, Alkalis.

RNase Decontamination

This can be performed using the following protocol:-

Clean the units with a mild detergent as described above.

Wash with 3% hydrogen peroxide (H2O2) for 10 minutes.

Rinsed with 0.1% DEPC- (diethyl pyrocarbonate) treated distilled water,

Caution:

DEPC is a suspected carcinogen. Always take the necessary precautions when using. RNaseZAP™ (Ambion) can also be used.

Please consult the instructions for use with acrylic gel tanks.

10  

 

Section 2

Operating Instructions

2.1) Setting up the omniPAGE Gel Tank

Note: Before setting up the Gel Tank please ensure that it has been properly cleaned and dried.

1.

Note the position of the lid on the unit. This shows the correct polarity and the correct orientation of the cables, black is negative and red positive.

2.

Remove the lid from the unit.

Note: If the lid is not removed, fitting the cables may result in un-tightening of the gold plug and damage the electrode.

3.

Screw the cables into the tapped holes as fully as possible so that there is no gap between the lid and the leading edge of the cable fitting.

4.

Refit the lid and

the unit is now ready to be used.

2.2) Gel casting for CVS10, VS10W, CVS20 and CVS30

2.2.1) Cleaning the Glass Plates

Clean a set of glass pl ates for each gel first with distilled water and then w ith

-

-

70 % ethanol.

One set of glass pl ates constitutes one notched glass plate and one plain glass plate with bonded spacers.

When using a trip le glass plate sandwich, two notched glass plates are required, one set of free spacers and a set of plain glass plates with bonded spacers. The plain glass plate is positioned outermost, then a notched glass plate, free spacers and second notched glass plate. Alternatively, accesso ry notch glass plates with bonded spacers are available.

Note: All glass plates, gel casting modules, casting base and accessories must be completely dry before the set – up.

Wet components are more likely to miss-align and cause leaks.

11  

2 .2.2) Glass cassette Assembly

A ssemble the glass plates so that the bottom of the glass plates and the

spacers are perfec tly aligned.

For triple plate sandwiches, the free spacers Need to be perfectly aligned which is best performe d using a small spacer or comb to push the spacers apart. Notched glass plates with bonded spacers do not need manual alignment.

 

N OTE: The glass plates with bonded spacers have an arrow in the top of the spacers which are slightly longer than the glass plate to indicate the top.

2 .2.3) Casting Stand Assembly

-

Position the Slab Gel Insert on a flat surface.

-

-

Insert the glass pl ates into the Slab Gel Insert between the pressure bar and the blue gasket.

The Slab Gel Insert contains pressure bars which impart even pressure onto the glass pl ates and allow even screw pressure transfer onto the sealing edge of the glass plate, ensuring complete sealing. Ensure that the pressure bars are adequately open for the thickness of spacer used. The ba r can be opened by loosening the screws or by sliding the clamps. When using a triple glass plate sandwich, the pressure bars will need to be in the completely open position.

12  

 

T hen fully tighten the pressure bar screws in the order top then bottom.

SCREW   VERSION SLIDING   CLAMP   VERSION  

Fully tighten the screw for the Mini vertical and the screws sequentially and in an even manner for the maxi vertical in the order middle two, top then bottom, making sure not to wobble the unit. When using the Slide Clamp

Mini version, simply slide both gates outwards until fully tightened. When only one gel is being run, the dummy plate must be used in the second position and fully tightened.

NOTE: At this stage, check that the bottom edges of the spacers and glass plates are perfectly aligned.

Position the Slab Gel Insert in the casting base such that the Cam pins have handles pointing downwards and are located in the insert holes. The top of the GRM may need to be pushed down very slightly to locate the cam pins.

SCREW   VERSION SLIDING   CLAMP   VERSION

With the cam pin handles facing directly downwards, turn the cam pins fully through 180

0 or until the insert has tightened onto the silicone mat.

13  

 

NOTE: It is best to turn the cams in opposite directions to each other.

Do not overturn as this will cause the glass plates to push upwards and the assembly will be more likely to leak. The unit is now ready for gel preparation and pouring

Always reverse the silicone mat after casting to avoid indentations from persisting. Never leave the casting up-stand with glass plates tightened into the casting base for long periods of time as this will also cause indentations in the silicone mat.

The slide clamp version CVS10 also includes screws. This system can be used either with the slide clamps or screws as preferred by the user.

For those that prefer to use the screws rather than clamps, the screws can be simply inserted into the screw holes. The clamps can be removed by placing each clamp in the fully open position and gently bending the clamp upwards from the slanted end. The holding pin will then slowly release and the clamp can be removed.

2.2.4) Gel Pouring

-

Casting a gel with stacking layer:-

1) Place a comb into the gel cassette assembly with any gel and mark the glass plate below the comb teeth. This is the reference level to which the resolving gel should be poured.

2) Prepare the resolving gel solution. Mix well and avoid generating air bubbles.

3) Fill the glass plates smoothly till the mark avoiding generating any air bubbles. Filling must be performed quickly before the TEMED causes the gel to become too viscous.

4) Overlay the gel extremely carefully with 1 ml of Isobutanol, Isopropanol or distilled water. When using distilled water extra care must be taken to ensure there is no mixing with the gel solution.

5) Let the resolving gel polymerize. Usually this takes around 15 to 30 minutes but this can vary due to the freshness of the reagents used. If polymerization is taken a lot longer than this, use fresher stock solutions or add more APS and TEMED.

14  

 

-

6) Prepare the stacking gel solution.

7) Before casting the stacking gel, insert a piece of filter paper to dry the area in between the glass plates above the resolving gel. Take care not to touch the surface of the gel.

8) Carefully pour the stacking gel solution, avoiding generating air bubbles.

9) Carefully insert the comb making sure that no air bubbles get trapped under the ends of the comb teeth as these will inhibit sample progression.

10) Allow the stacking gel polymerize.

11) Once the gel is polymerized it is ready for the electrophoresis run.

Casting a gel without stacking layer:-

1) Prepare the resolving gel solution. Mix well and avoid generating air bubbles.

2) Pour the solution smoothly into the glass plates avoiding any air bubbles until the top of the notched glass plate is reached.

3) Carefully insert the comb making sure that no air bubbles get trapped under the ends of the comb teeth as these will inhibit sample progression.

4) Let the gel polymerize. Usually this takes from 15 to 30 minutes but this can vary due to the freshness of the reagents used. If polymerization is taken a lot longer than this, use fresher stock solutions or add more APS and TEMED.

5) Once the gel is polymerized it is ready for the electrophoresis run.

-

Using Precast Gels:-

1) omniPAGE mini is compatible with all the precast gels available in the market.

2) Simply remove the precast gel from the storage pouch.

3) Gently remove the comb.

4) Keep the Inner module upstand on a flat surface and place the precast gel between the pressure bar and the blue gasket.

15  

 

2.3) omniPAGE module assembly and Sample loading

1) If desired, fit the cooling pack(s) into the end of the tank. These should be pre-frozen and fitted with the longest side positioned sideways with the end(s) of the tank and pressed into the recess. Or these can be fitted down the front of the tank.

Note: NEVER FIT THESE UNDERNEATH THE MODULE IN THE BOTTOM OF

THE TANK AS THIS WILL PREVENT THE FLOW OF CURRENT THROUGH THE

GEL AND CAUSE SLOW RUNS AND OVER-HEATING.

Note one pack is supplied as standard. Additional packs can be purchased.

2) Transfer the Inner gel module containing cast gels into the main tank in the correct orientation as indicated - +ve on the module aligned with +ve on the tank, -ve on the module aligned with –ve on the tank.

3) Fill the outer tank with 1 x reservoir buffer. See Page 22 for recommended running buffer solution.

Table 7

shows the volume of buffer required.

4) Load the samples into the wells using a pipette tip taking care not to damage the wells or induce any air bubbles.

5) Fill any unused wells with 1 X sample buffer.

6) It is a good idea to note the orientation and order the samples were loaded in. This can be done by noting which samples were loaded adjacent to each electrode.

2.4) Gel Running:-

1)

Fit the lid and connect to a power supply.

2) Consult Table 8 for details on recommended power supply voltage settings.

2.5) Gel Removal:-

1) Turn the power supply off when the loading dye reaches the bottom of the gel, sooner if your proteins are below 4Kd in size.

2) Remove the gel running module, first emptying the inner buffer into the main tank. Buffer can be re-used but this may affect run quality if continued.

3) Unscrew the glass plates with the Screw version. To open the sliding door version insert the CVS10KEY into the recess arch of the clamping door. Twist key applying pressure to both the clamping door and the CVS10D side cheek.

16  

The door will now click open. Repeat this process until you have opened both the doors.

4) Remove the glass plates. Then using CSLKEY separate notched and the plain glass plates. Place the wedged end of the key between the two plates and gently twist until the plates pull apart. The gel will usually stick to one of the plates and can be removed by first soaking in buffer and then gently lifting with a spatula.

5) The gel is now ready to be stained with Coomassie or silver stain or the proteins in the gel can be transferred to a membrane by electroblotting for specific band identification and further analysis.

3.1) Gel Selection

Care should be taken when selecting the pore size of the gel to be used.

The pore size or % of gel determines the resolving ability given different sizes of protein.

See Table 1 below explaining which percentage of gel to use to separate the sizes of proteins indicated.

Table 1

Acrylamide Percentage

5 %

7.5 %

10 %

12%

15 %

17.5%

Separating Resolution

60 - 220 KD

30 - 120 KD

20 - 75 KD

17 – 65 KD

15 -45 KD

12 – 30 KD

Section 3

Gel Preparation

17  

 

3.2) Volumes required per gel

Table 2 & 3

below shows the total amount of gel solution required.

Table 2 omniPAGE Mini

VS10D, VS10DSYS, CVS10D, CVS10DSYS omniPAGE Mini Wide

VS10WD, VS10WDSYS

Number of gels

Single – one gel, one dummy plate

Double – two gels

Using a Triple Plate sandwich – four gels

Gel

Thickness

(mm)

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

Volume

(ml)

3.8

7.5

11.3

15.0

7.5

15.0

22.5

30.0

10.0

30.0

45.0

60.0

Number of gels

Single – one gel, one dummy plate

Double – two gels

Using a Triple Plate sandwich – four gels

Gel

Thickness

(mm)

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

Volume

(ml)

8.75

17.5

26.3

35.0

17.5

35.0

52.5

70.0

35.0

70.0

105.0

140.0

Table 3 omniPAGE Maxi – VS20D, VS20DSYS

Number of gels

Single – one gel, one dummy plate

Double – two gels

Using a Triple Plate sandwich – four gels

Gel

Thickness

(mm)

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

O.5

1.0

1.5

2.0

Volume

(ml)

17.5

35.0

52.5

70.0

35.0

70.0

105.0

140.0

70.0

140.0

210.0

280.0

omniPAGE - VS30

For different thicknesses of gel, multiple the amounts by the spacer thickness. * multiply by 1.5 for

VS30 gels

18  

 

3.3) Gel Preparation

Prepare gel solutions as per tables below. These give the volumes of solutions from the standard stock solutions. These should be gently mixed avoiding generation of bubbles which will inhibit polymerization by removing free radicals.

Table 4:

Preparation of the separating gel solution for two 10 x 10cm (C)VS10D gels using 1 mm spacers.

Solution

Distilled Water

5 %

8.7ml

30 % Stock Acrylamide

Solution

2.5ml

7.5%

7.5ml

10 %

6.3ml

3.75ml 5ml

12%

5.25ml

6ml

15 % 17.5%

3.75ml 2.5ml

7.5ml 8.75ml

4 X Resolving Tris

Solution

10 % Ammonium

Persulphate

150µl 150µl 150µl 150µl 150µl 150µl

Table 5:

Preparation of the separating gel solution for two 20 x 20cm VS20D gels using

1 mm spacers. Divide by two for VS10W gels. Multiply by 1.5 for VS30 gels

Solution 5 % 7.5% 10 % 12% 15 % 17.5%

Distilled Water

30 % Stock

Acrylamide Solution

41ml 35.25ml 29.6ml 24.7ml 17.6ml

11.7ml 17.6ml

11.7ml

23.5ml 28.2ml 35.25ml 41.1ml

4 X Resolving Tris

Solution

10 % Ammonium

Persulphate

700µl 700µl 700µl 700µl 700µl 700µl

 

Add 15µl of TEMED to the resolving gel solution for (C)VS10D sized gels, 35 µl for

VS10W, 70µl for VS20D and 105 µl for VS30D gels and mix well but avoid generating air bubbles.

19  

 

3.4) Stacking Gel Preparation

Prepare the stacking gel using Table 6 below as a guide.

Table 6:

Distilled Water

30 % Stock Acrylamide

Solution

4 X Stacking Gel Tris

Solution

10 % Ammonium

Persulphate

4.2ml 8.4ml 16.8ml 25.2ml

0.65ml 1.3ml 2.6ml 3.9ml

1.6ml 3.2ml 6.4ml 9.6ml

Add 6.7µl of TEMED to the stacking gel solution for VS10 gels. For VS10W gels add

13.4 µl, for VS20 gels add 26.8µl and 40.2 µl for VS30 gels. Mix well. Use a Pasteur pipette to fill the glass plates up to the top with stacking gel solution.

3.5) Buffer Volume

Table 7

Buffer Volume

Minimum

– Inner tank is filled to above the wells. Outer

Tank is filled to just flood the bottom of the glass plates.

Cooling potential is at a minimum which may affect resolution.

(C)VS10D VS20D,

VS10WD

250ml

500ml

Maximum

– Inner tank is filled to above the wells. Outer

Tank is filled to the maximum fill line. Cooling is high offering good resolution of samples.

1200ml

2.8 Litres

Using the cooling packs

– Inner tank is filled to above the wells. Cooling packs are inserted behind the gels. Outer

Tank is filled to the maximum fill line. Cooling is at a maximum.

1000ml

2.3 Litres

VS30D

1.2Litres

1.8 Litres

5.6 Litres

8.4 Litres

4.6Litres

6.9 Litres

20  

3.6) Gel Running Conditions

Table 8

Recommended Voltages and Resultant Current for

1mm thick, 12% gels.

One gel

Two gels

Three gels

Four gels

(C)VS10D,

VS10WD

VS20D

VS30D

90-225V

20-45mA

90-225V

40-90mA

90-225V

60-135mA

90-225V

80-180mA

120-250V

20-45mA

120-250V

40-90mA

120-250V

60-135mA

120-250V

80-180mA

3.7) Preparation of denatured protein samples for loading:

The instructions given below are for denatured samples. For Native samples, please consult a laboratory handbook.

1. Prepare the protein samples for loading. The volume of sample depends on the capacity of the wells (See Comb specifications pages 22 and 23).

2. Using a 0.5 ml micro-centrifuge tube or other convenient receptacle, combine the protein sample and 4 X sample buffer. It is always advisable to use protein markers in one of the end lanes to indicate sizes of bands. These should be prepared according to the manufacturer’s instructions.

3. Heat the samples in a water bath or heating block for 2 minutes to denature the samples.

4. Centrifuge the samples in a micro-centrifuge for 20 seconds at 12,000 rpm. The protein samples are now ready to load.

21  

 

Section 4

Stock Solutions

Stock Solutions for SDS PAGE gels:-

Stock 30% Acrylamide Gel Solution:-

30.0 g acrylamide

0.8 g methylene bisacrylamide

Distilled Water to 100ml

Stock 4 X Resolving Gel Tris (1.5 M Tris

.

HCl pH8.8, 0.4 % SDS)

To 110ml Distilled Water add 36.4 g of Tris base

Add 8ml of 10 % SDS

Adjust pH to 8.8 with 1N HCl

Adjust the final volume to 200ml with Distilled Water.

Stock 4 X Stacking Tris (0.5 M Tris

.

HCL pH6.8, 0.4 % SDS)

To 110ml Distilled Water add 12.12 g of Tris base

Add 8ml of 10 % SDS

Adjust pH to 6.8 with 1N HCl

Add Distilled Water to a final volume of 200ml

Stock 4 X Tris-glycine tank buffer - SDS

36 g Tris base

172.8 g glycine

Distilled Water to 3 L

1 x Tris-glycine tank buffer - SDS

750ml of 4 X Tris-glycine reservoir buffer - SDS

30ml of 10 % SDS

Distilled Water to 3L

10 % AP (ammonium persulphate solution)

0.1 g ammonium persulphate

1ml Distilled Water

22  

 

TEMED

Stock 4 X Sample Buffer

4ml glycerol

2ml 2-mercaptoethanol

1.2 g SDS

5ml 4 X Stacking Tris

0.03 g Bromophenol blue

Aliquot into 1.5ml micro centrifuge tubes. Store at -20°C.

Section 5 Ordering Information

5.1) omniPAGE Mini

 

Catalogue Number Description

CVS10D

CVS10DSYS

CVS10PRE

CVS10DSYS-CU omniPAGE Mini, 10 x 10cm Dual, 2 sets of Glass Plates, 1mm thick bonded

Spacers, 2 x 12 sample, 1mm thick combs. CLAMP VERSION omniPAGE Mini, 10 x 10cm Dual, 2 sets of Glass Plates, 1mm thick bonded

Spacers, 2 x 12 sample, 1mm thick combs including caster. CLAMP VERSION omniPAGE Mini, 10 x 10cm Dual. No accessories. CLAMP VERSION omniPAGE Mini, 10 x 10cm Dual, 2 sets of Glass Plates, 1mm thick bonded

Spacers, 2 x 12 sample, 1mm thick combs including caster. CLAMP VERSION,

External casting upstand omniPAGE Mini Accessories

Catalogue Number Description

CVS10EXCASTER

CVS10EXCASTERSYS

VS10DCAST

VS10DCASTM

VS10DIRM*

CVS10DIRM†

VS10ICB

VS10-X-LG

VS10NG

VS10PG

VS10NGS0.75

VS10PGS0.75

External Casting Stand - No Casting Base

External Casting System - Upstand+ Base

10 x 10cm Casting Base

Replacement Silicone Mat for 10 x 10cm Casting Base

Inner Running Module

Inner Running Module

Mini Cooling Pack

Loading Guides x = sample number

10 x 10cm Notched Glass Plates 2mm thick (pk/2)

10 x 10cm Plain Glass Plates 2mm thick (pk/2)

10 x 10cm Notched Glass Plates with 0.75mm Bonded Spacers (pk/2)

10 x 10cm Plain Glass Plates with 0.75mm Bonded Spacers (pk/2)

23  

VS10NGS1

VS10PGS1

VS10NGS1.5

VS10PGS1.5

VS10NGS2

VS10PGS2

VS10DP

VS10S0.75

VS10S1

VS10S1.5

VS10S2

RPW-0.2

10 x 10cm Notched Glass Plates with 1mm Bonded Spacers (pk/2)

10 x 10cm Plain Glass Plates with 1mm Bonded Spacers (pk/2)

10 x 10cm Notched Glass Plates with 1.5mm Bonded Spacers (pk/2)

10 x 10cm Plain Glass Plates with 1.5mm Bonded Spacers (pk/2)

10 x 10cm Notched Glass Plates with 2mm Bonded Spacers (pk/2)

10 x 10cm Plain Glass Plates with 2mm Bonded Spacers (pk/2)

Dummy Plate, 10 x 10cm

10cm Spacers - 0.75mm (pk/2)

10cm Spacers - 1mm thick (pk/2)

10cm Spacers - 1.5mm thick (pk/2)

10cm Spacers - 2mm thick (pk/2)

Replacement Platinum Wire - 0.2mm, 50cm

Multiple Minigel Casting

Catalogue Number Description

CSL-6CAST

CSL-12CAST

CSL-24CAST

6 gel caster for 8 x 10cm or 10 x 10cm gels

12 gel caster for 8 x 10cm or 10 x 10cm gels

24 gel caster for 8 x 10cm or 10 x 10cm gels

-

Variety of combs available with VS10D

MC Denotes Multi Channel Pipette compatible.

Code Description

Sample Volume

µl for a 5mm thick gel

VS20-1-0.75

VS20-5-0.75

VS20-10-0.75

VS20-18-0.75MC

VS20-24-0.75

VS20-30-0.75

VS20-36-0.75MC

VS20-48-0.75

Comb 1 Prep, 1 Marker, 0.75mm thick

Comb 5 sample, 0.75mm thick

Comb 10 sample, 0.75mm thick

Comb 18 sample MC, 0.75mm thick

Comb 24 sample, 0.75mm thick

Comb 30 sample, 0.75mm thick

Comb 36 sample MC, 0.75mm thick

Comb 48 sample, 0.75mm thick

40

30

25

20

15

1100

160

80

24  

 

 

VS20-36-1.5MC

VS20-48-1.5

VS20-1-2

VS20-5-2

VS20-10-2

VS20-18-2MC

VS20-24-2

VS20-30-2

VS20-36-2MC

VS20-48-2

VS20-1-1

VS20-5-1

VS20-10-1

VS20-18-1MC

VS20-24-1

VS20-30-1

VS20-36-1MC

VS20-48-1

VS20-1-1.5

VS20-5-1.5

VS20-10-1.5

VS20-18-1.5MC

VS20-24-1.5

VS20-30-1.5

Comb 1 Prep, 1 Marker, 1mm thick

Comb 5 sample, 1mm thick

Comb 10 sample, 1mm thick

Comb 18 sample, 1mm thick

Comb 24 sample, 1mm thick

Comb 30 sample, 1mm thick

Comb 36 sample MC, 1mm thick

Comb 48 sample, 1mm thick

Comb 1 Prep, 1 Marker, 1.5mm thick

Comb 5 sample, 1.5mm thick

Comb 10 sample, 1.5mm thick

Comb 18 sample, 1.5mm thick

Comb 24 sample, 1.5mm thick

Comb 30 sample, 1.5mm thick

Comb 36 sample MC, 1.5mm thick

Comb 48 sample, 1.5mm thick

Comb 1 Prep, 1 Marker, 2mm thick

Comb 5 sample, 2mm thick

Comb 10 sample, 2mm thick

Comb 18 sample, 2mm thick

Comb 24 sample, 2mm thick

Comb 30 sample, 2mm thick

Comb 36 sample MC, 2mm thick

Comb 48 sample, 2mm thick

25  

40

30

3000

400

200

100

80

70

50

40

25

20

2200

320

160

80

60

50

1500

200

100

50

40

35

 

5.2) omniPAGE Mini Wide

Catalogue Number Description

VS10WD

VS10WDSYS

VS10WDSYS-CU omniPAGE Mini Wide, 20 x 10cm Dual, 2 sets of Glass Plates with 1mm thick bonded Spacers, 2 x 24 sample, 1mm thick combs, cooling pack omniPAGE Mini Wide, 20 x 10cm Dual, 2 sets of Glass Plates with 1mm thick bonded Spacers, 2 x 24 sample, 1mm thick combs, cooling pack including caster omniPAGE Mini Wide, 20 x 10cm Dual, 2 sets of Glass Plates, 1mm thick bonded Spacers, 2 x 24 sample, 1mm thick combs including caster, External casting upstand

Accessories

VS10WEXCASTER

VS20CAST

VS20DCASTM

VS10WDIRM

VS20-x -LG

VS10WNG

VS10WPG

VS10WNGS0.75

VS10WPGSO.75

VS10WNGS1

VS10WPGS1

VS10WPGS1.5

VS10WPGS2

VS10WDP

RPW-0.2

VS20ICB

Catalogue Number Description

VS10W External Casting Stand - No Casting Base

20 x 10cm Casting Base

Replacement Silicone Mat for 20 x 10cm Casting Base

Inner Running Module

Loading guides for omniPAGE mini combs, x = comb well number

20 x 10cm Notched Glass Plates 4mm thick (pk/2)

20 x 10cm Plain Glass Plates 4mm thick (pk/2)

20 x 10cm Notched Glass Plates with 0.75mm Bonded Spacers (pk/2)

20 x 10cm Plain Glass Plates with 0.75mm Bonded Spacers (pk/2)

20 x 10cm Notched Glass Plates with 1mm Bonded Spacers (pk/2)

20 x 10cm Plain Glass Plates with 1mm Bonded Spacers (pk/2)

20 x 10cm Plain Glass Plates with 1.5mm Bonded Spacers (pk/2)

20 x 10cm Plain Glass Plates with 2mm Bonded Spacers (pk/2)

Dummy Plate, 20 x 10cm

Replacement Platinum Wire - 0.2mm, 50cm

Maxi Cooling Pack

5.3) omniPAGE Maxi

Catalogue Number

VS20D

VS20DSYS

Description omniPAGE Maxi, 20 x 20cm Dual with Glass Plates with bonded 1mm thick spacers, 2x 24 sample combs, cooling pack, dummy plate omniPAGE Maxi, 20 x 20cm Dual with Glass Plates with bonded 1mm thick

26  

 

VS20DSYS-CU spacers, 2x 24 sample combs, cooling pack, dummy plate and Casting Base omniPAGE Maxi, 20 x 20cm Dual, 2 sets of Glass Plates, 1mm thick bonded

Spacers, 2 x 24 sample, 1mm thick combs including caster, External casting upstand

Accessories

Catalogue Number

VS20WEXCASTER

VS20DCAST

VS20DCASTM

VS20DIRM

VS20ICB

VS20-x -LG

VS20NG

VS20PG

VS20NGS0.75

VS20PGS0.75

VS20NGS1

VS20PGS1

VS20PGS1.5

VS20PGS2

VS20DP

VS20S0.75

VS20S1

VS20S1.5

VS20S2

RPW-0.2100

Description

VS20 External Casting Stand - No Casting Base

20 x 20cm Dual Casting Base

Replacement Silicone Mat for 20 x 20cm Casting Base

Inner Running Module

Maxi Cooling Pack

Loading guides for omniPAGE maxi combs, x = comb well number

20 x 20cm Notched Glass Plates 4mm thick (pk/2)

20 x 20cm Plain Glass Plates 4mm thick (pk/2)

20 x 20cm Notched Glass Plates with 0.75mm Bonded Spacers (pk/2)

20 x 20cm Plain Glass Plates with 0.75mm Bonded Spacers (pk/2)

20 x 20cm Notched Glass Plates with 1mm Bonded Spacers (pk/2)

20 x 20cm Plain Glass Plates with 1mm Bonded Spacers (pk/2)

20 x 20cm Plain Glass Plates with 1.5mm Bonded Spacers (pk/2)

20 x 20cm Plain Glass Plates with 2mm Bonded Spacers (pk/2)

Dummy Plate, 20 x 20cm

20cm Spacers - 0.75mm (pk/2)

20cm Spacers - 1mm thick (pk/2)

20cm Spacers - 1.5mm thick (pk/2)

20cm Spacers - 2mm thick (pk/2)

Replacement Platinum Wire - 0.2mm, 100cm

5.4) Variety of combs available with VS20D, VS10WD:-

Code Description

VS20-1-0.75

VS20-5-0.75

Comb 1 Prep, 1 Marker, 0.75mm thick

Comb 5 sample, 0.75mm thick

Sample Volume

µl for a 5mm thick gel

1100

160

27  

 

VS20-1-1.5

VS20-5-1.5

VS20-10-1.5

VS20-18-1.5MC

VS20-24-1.5

VS20-30-1.5

VS20-36-1.5MC

VS20-48-1.5

VS20-1-2

VS20-5-2

VS20-10-2

VS20-18-2MC

VS20-24-2

VS20-10-0.75

VS20-18-0.75MC

VS20-24-0.75

VS20-30-0.75

VS20-36-0.75MC

VS20-48-0.75

VS20-1-1

VS20-5-1

VS20-10-1

VS20-18-1MC

VS20-24-1

VS20-30-1

VS20-36-1MC

VS20-48-1

Comb 10 sample, 0.75mm thick

Comb 18 sample MC, 0.75mm thick

Comb 24 sample, 0.75mm thick

Comb 30 sample, 0.75mm thick

Comb 36 sample MC, 0.75mm thick

Comb 48 sample, 0.75mm thick

Comb 1 Prep, 1 Marker, 1mm thick

Comb 5 sample, 1mm thick

Comb 10 sample, 1mm thick

Comb 18 sample, 1mm thick

Comb 24 sample, 1mm thick

Comb 30 sample, 1mm thick

Comb 36 sample MC, 1mm thick

Comb 48 sample, 1mm thick

Comb 1 Prep, 1 Marker, 1.5mm thick

Comb 5 sample, 1.5mm thick

Comb 10 sample, 1.5mm thick

Comb 18 sample, 1.5mm thick

Comb 24 sample, 1.5mm thick

Comb 30 sample, 1.5mm thick

Comb 36 sample MC, 1.5mm thick

Comb 48 sample, 1.5mm thick

Comb 1 Prep, 1 Marker, 2mm thick

Comb 5 sample, 2mm thick

Comb 10 sample, 2mm thick

Comb 18 sample, 2mm thick

Comb 24 sample, 2mm thick

28  

3000

400

200

100

80

80

60

50

2200

320

160

40

30

50

40

35

1500

200

100

25

20

80

40

30

25

20

15

 

VS20-30-2

VS20-36-2MC

VS20-48-2

Comb 30 sample, 2mm thick

Comb 36 sample MC, 2mm thick

Comb 48 sample, 2mm thick

5.5) omniPAGE VS30

70

50

40

Catalogue Number Description

VS30D omniPAGE Maxi Plus, 30 x 22cm Dual with Glass Plates with bonded 1.5mm spacers, 2 x 28 sample combs, 2 x 2-D combs, cooling pack, dummy plate

Accessories

VS30DSYS

VS30BI

VS30DCAST

VS30DCASTM

VS30DIRM

VS30ICB

VS30-x -LG

VS30NG

VS30PG

VS30NGS0.75

VS30PGS0.75

VS30NGS1

VS30PGS1

VS30PGS1.5

VS30PGS2

VS30DP

VS30S0.75

VS30S1

VS30S1.5

VS30S2

RPW-0.2100

Catalogue Number Description

VS30D with Casting Base omniPAGE VS30 Blot Maxi Insert - includes 3 cassettes and 6 fibre pads.

30 x 22cm Dual Casting Base

Replacement Silicone Mat for 30 x 22cm Casting Base

Inner Running Module

Maxi Cooling Pack

Loading guides for omniPAGE maxi combs, x = comb well number

30 x 22cm Notched Glass Plates 4mm thick (pk/2)

30 x 22cm Plain Glass Plates 4mm thick (pk/2)

30 x 22cm Notched Glass Plates with 0.75mm Bonded Spacers (pk/2)

30 x 22cm Plain Glass Plates with 0.75mm Bonded Spacers (pk/2)

30 x 22cm Notched Glass Plates with 1mm Bonded Spacers (pk/2)

30 x 22cm Plain Glass Plates with 1mm Bonded Spacers (pk/2)

30 x 22cm Plain Glass Plates with 1.5mm Bonded Spacers (pk/2)

30 x 22cm Plain Glass Plates with 2mm Bonded Spacers (pk/2)

Dummy Plate, 30 x 22cm

22cm Spacers - 0.75mm (pk/2)

22cm Spacers - 1mm thick (pk/2)

22cm Spacers - 1.5mm thick (pk/2)

22cm Spacers - 2mm thick (pk/2)

Replacement Platinum Wire - 0.2mm, 100cm

29  

-

Variety of combs available with VS30D:-

Code Description

 

VS30-1-1

VS30-2-1

VS30-4-1

VS30-28-1MC

VS30-56-1MC

VS30-75-1.5

VS30-1-1.5

VS30-2-1.5

VS30-4-1.5

VS30-28-1.5MC

VS30-56-1.5MC

VS30-75-1.5

Comb 1 Prep, 1 Marker, 1mm thick

Comb 2 sample, 1mm thick

Comb 4 sample, 1mm thick

Comb 28 sample, 1mm thick MC compatible

Comb 56 sample, 1mm thick MC compatible

Comb 75 sample, 1mm thick

Comb 1 Prep, 1 Marker, 1.5mm thick

Comb 2 sample, 1.5mm thick

Comb 4 sample, 1.5mm thick

Comb 28 sample, 1.5mm thick MC compatible

Comb 56 sample, 1.5mm thick MC compatible

Comb 75 sample, 1.5mm thick

30  

Sample Volume

µl for a 5mm thick gel

25

3375

1680

825

120

60

37

2250

1125

550

80

40

 

Section 6 Warranty

The Cleaver Scientific Ltd. (CSL) Electrophoresis units have a warranty against manufacturing and material faults of twelve months from date of customer receipt.

If any defects occur during this warranty period, CSL will repair or replace the defective parts free of charge.

This warranty does not cover defects occurring by accident or misuse or defects caused by improper operation.

Units where repair or modification has been performed by anyone other than CSL or an appointed distributor or representative are no longer under warranty from the time the unit was modified.

Units which have accessories or repaired parts not supplied by CSL or its associated distributors have invalidated warranty.

CSL cannot repair or replace free of charge units where improper solutions or chemicals have been used. For a list of these please see the Care and Maintenance subsection.

If a problem does occur then please contact your supplier or CSL on:-

Cleaver Scientific Ltd.

Unit 4 Triton Park

Swift Valley

Brownsover Road

Rugby

CV21 1SG

Tel: +44 (0)1788 565300

Fax: +44 (0)1788 552822

Email: [email protected]

31  

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